BackgroundOur previous strain‐specific systematic review (SR) showed that Lactobacillus reuteri (LR) DSM 17938 reduces necrotizing enterocolitis (NEC), late onset sepsis (LOS), and time to full feeds (TFF) in preterm infants. Considering progress in the field over last six years, we aimed to update our SR.MethodsSR of randomized controlled trials (RCTs) and non‐RCTs was conducted. MEDLINE, EMBASE, EMCARE, Cochrane CENTRAL and grey literature databases were searched in June 2023. Primary outcomes: TFF, NEC ≥Stage II, LOS, and all‐cause mortality. Meta‐analysis was performed using random effects model. Certainty of Evidence (CoE) was summarized using GRADE guidelines. Trial Sequential Analysis (TSA) was applied for the outcome of NEC in RCTs.ResultsTwelve RCTs (n=2284) and four non‐RCTs (n=1616) were included. Of them, six RCTs and three non‐RCTs were new. Meta‐analysis of RCTs showed LR significantly reduced TFF [MD: ‐2.70 (95% CI: ‐4.90 to ‐1.31) days; p=0.0001), NEC ≥stage II [RR: 0.57 (95% CI: 0.37‐ 0.87); p=0.009, 8 RCTs) and LOS [RR: 0.72 (95% CI: 0.54‐0.97); p=0.03). There was no significant reduction in mortality [RR: 0.76 (95% CI: 0.54‐1.06); p=0.10). TSA showed diversity adjusted required information size (DARIS) as 3624 for NEC. Overall CoE: ‘very low’. Meta‐analysis of four non‐RCTs showed LR significantly reduced NEC [OR: 0.34 (95% CI: 0.15‐0.77; p=0.01) but not LOS. LR had no adverse effects.ConclusionsVery low CoE suggests that LR DSM 17938 may reduce risk of NEC, LOS, and shorten TFF in preterm infants. Additional RCTs are required to increase sample size and CoE.This article is protected by copyright. All rights reserved.